KeyGene (Netherlands)

Apomixis, clonal reproduction through seed, is divided into sporophytic (nucellar embryony) and gametophytic apomixis. This book focuses on the latter type. Gametophytic apomixis, while rare, is taxonomically widespread, notably among grasses and Asteraceae. Detection methods have advanced significantly, employing flow cytometry and advanced microscopy techniques. The genetic, molecular, and developmental mechanisms behind apomixis involve modifications preventing meiosis and enabling parthenogenesis. Understanding these mechanisms and applying approaches such as forward genetics and mutagenesis holds an important potential for agricultural innovation. Numerous genetic, ecological, and evolutionary questions remain open, highlighting apomixis as an important field for ongoing scientific research.

Management of the phloem-feeding pest insect Bemisia tabaci (whitefly) is difficult due to its short generation time and large number of offspring. Several whitefly-resistant wild tomato accessions have been identified, with the resistance attributed to specific defense metabolites in glandular trichomes. Interestingly, we found that on Solanum chmielewskii LA1840, which lacks trichome-based resistance, nymphal development is delayed and decreased compared with a cultivated tomato. Here, we show that the resistance observed in LA1840 is based on a mobile factor in the vasculature, the site of interaction for nymphs during feeding. The putative compound responsible for the resistance apparently passed the graft junction from an LA1840 rootstock to an otherwise susceptible cultivar scion. After untargeted metabolomics on the phloem collected from the wild accessions, a Random Forest algorithm predicted riboflavin to be linked to the resistance phenotype. The resistant genotypes indeed exhibit increased riboflavin levels in leaves compared with susceptible plants. The effect of elevated riboflavin levels on whitefly nymph development was validated through feeding riboflavin to susceptible plants. Our results highlight the power of natural variation in metabolites and vasculature-based resistance mechanisms for the development of sustainable whitefly management.

Nucleotide-binding leucine-rich repeat receptor (NLR) genes encode a pivotal class of plant immune receptors. However, their rampant duplication and loss have made inferring their genomic evolutionary trajectory difficult, exemplified by the loss of TNL family genes in monocots. In this study, we introduce a novel classification system for angiosperm NLR genes, grounded in network analysis of microsynteny information. This refined classification categorizes these genes into five classes: CNL_A, CNL_B, CNL_C, TNL, and RNL. Compared to the previous classification, we further subdivided CNLs into three subclasses. The credibility of this classification is supported by phylogenetic analysis and examination of protein domain structures. Importantly, this classification enabled a model to explain the extinction of TNL genes in monocots. Compelling microsynteny evidence underscores this revelation, indicating a clear synteny correspondence between the non-TNLs in monocots and the extinct TNL subclass. Our study provides crucial insights into the genomic origin and divergence of plant NLR subfamilies, unveiling the malleability-driven journey that has shaped the functionality and diversity of plant NLR genes.

Hybrid seed failure arising from wide crosses between plant species is a recurring obstacle in plant breeding, impeding the transfer of desirable traits. This postzygotic reproductive barrier primarily occurs in the endosperm, a tissue that nourishes the embryo and functions similarly to the placenta in mammals. We found that incompatible seeds show a loss of DNA methylation and chromatin condensation in the endosperm, similar to seeds lacking maternal RNA polymerase IV activity. This similarity is linked to a decrease in small interfering RNAs in the endosperm (sirenRNAs), maternal RNA polymerase IV-dependent short interfering RNAs that regulate DNA methylation. Several AGAMOUS-like MADS-box transcription factor genes (AGLs), key regulators of endosperm development, are targeted by sirenRNAs in cis and in trans. This finding aligns with the enrichment of AGL target genes among deregulated genes. We propose that hybrid seed failure results from reduced maternal sirenRNAs combined with increased AGL expression, leading to abnormal gene regulation in the endosperm.

Abstract Improving traits by breeding involves compromising between desired traits and possible undesired side effects. Often encountered examples include pathogen resistance versus yield, shelf life time versus fruit quality, and seed number versus seed weight. Genome editing can be used to reduce the effect of some of these tradeoffs. Different genetic reasons underlying a tradeoff require different approaches: important to note is whether a detrimental effect is caused by a unique gene, or several analogs/ homologs, because the strategy needs to be adopted accordingly. Unique genes, for example, can be substituted by analogs, and homologs have the advantage that, amongst the available options, the gene causing the fewest pleiotropic effects can be altered in its activity. When the detrimental effect of a tradeoff is caused by two genetically linked genes, this can lead to linkage drag. To break this type of tradeoff genome editing can be used to force a crossover event. Overcoming a tradeoff can generate a new one, but can nevertheless result in an improved crop variety.

Accurate taxonomic classification of samples from infected host material is essential for disease diagnostics and genome analyses. Despite the importance, diagnosis of fungal pathogens causing banana leaf diseases remains challenging. Foliar diseases of bananas are mainly caused by 3 Pseudocercospora species, of which the most predominant causal agent is Pseudocercospora fijiensis. Here, we sequenced and assembled four fungal isolates obtained from necrotic banana leaves in Bohol (Philippines) and obtained a high-quality genome assembly for one of these isolates. The samples were initially identified as P. fijiensis using PCR diagnostics; however, the assembly size was consistently 30 Mb smaller than expected. Based on the internal transcribed spacer (ITS) sequences, we identified the samples as Zasmidium syzygii (98.7% identity). The high-quality Zasmidium syzygii assembly is 42.5 Mb in size, comprising 16 contigs, of which 11 are most likely complete chromosomes. The genome contains 98.6% of the expected single-copy BUSCO genes and contains 14,789 genes and 10.3% repeats. The 3 short-read assemblies are less continuous but have similar genome sizes (40.4-42.4 Mb) and contain between 96.5 and 98.4% BUSCO genes. All 4 isolates have identical ITS sequences and are distinct from Zasmidium isolates that were previously sampled from banana leaves. We thus report the first continuous genome assembly of a member of the Zasmidium genus, forming an essential resource for further analysis to enhance our understanding of the diversity of pathogenic fungal isolates as well as fungal diversity.

Abstract Accurate taxonomic classification of samples from infected host material is essential for disease diagnostics and genome analyses. Despite the importance, diagnosis of fungal pathogens causing banana leaf diseases remains challenging. Foliar diseases of bananas are mainly caused by threePseudocercosporaspecies, of which the most predominant causal agent isP. fijiensis. Here, we sequenced and assembled four fungal isolates obtained from necrotic banana leaves in Bohol (Philippines) and obtained a high-quality genome assembly for one of these isolates. The samples were initially identified asP. fijiensisusing PCR diagnostics, however, the assembly size was consistently 30 Mb smaller than expected. Based on the ITS sequences, we identified the samples asZasmidium syzygii(98.7% identity). The high-qualityZasmidium syzygiiassembly is 42.5 Mb in size, comprising 16 contigs, of which 11 are complete. The genome contains 98.6% of the expected single-copy BUSCO genes and contains 14,789 genes and 10.3% repeats. The three short-read assemblies are less continuous but have similar genome sizes (40.4 – 42.4 Mb) and contain between 96.5% and 98.4% BUSCO genes. All four isolates have identical ITS sequences and are distinct fromZasmidiumisolates that were previously sampled from banana leaves. We thus report the first continuous genome assembly of a member of theZasmidiumgenus, forming an essential resource for further analysis to enhance our understanding of the diversity of pathogenic fungal isolates as well as fungal diversity.

Bananas are major agricultural commodities in Cuba. One of the main constraints of banana production worldwide is Fusarium wilt of banana. Recent outbreaks in Colombia, Perú, and Venezuela have raised widespread concern in Latin America due to the potential devastating impact on the sustainability of banana production, food security, and livelihoods of millions of people in the region. Here, we phenotyped 18 important Cuban banana and plantain varieties with two Fusarium strains-Tropical Race 4 (TR4) and Race 1-under greenhouse conditions. These varieties represent 72.8% of the national banana acreage in Cuba and are also widely distributed in Latin America and the Caribbean region. A broad range of disease responses from resistant to very susceptible was observed against Race 1. On the contrary, not a single banana variety was resistant to TR4. These results underscore that TR4 potentially threatens nearly 56% of the contemporary Cuban banana production area, which is planted with susceptible and very susceptible varieties, and call for a preemptive evaluation of new varieties obtained in the national breeding program and the strengthening of quarantine measures to prevent the introduction of TR4 into the country.

In this paper we show that metabolic engineering in Cucurbita pepo hairy roots can be used to both effectively increase and modify cucurbitacins. Cucurbitacins are highly-oxygenated triterpenoids originally described in the Cucurbitaceae family, but have since been found in 15 taxonomically distant plant families. Cucurbitacin B, D, E and I are the most widespread amongst the Cucurbitaceae and they have both important biological and pharmacological activities. In this study C. pepo hairy roots were used as a platform to boost production and alter the structures of the afore mentioned cucurbitacins by metabolic engineering to potentially provide new or more desirable bioactivities. We report that the ability to induce cucurbitacin biosynthesis by basic Helix-Loop-Helix transcription factors is partially conserved within the Cucurbitaceae and therefore can potentially be used as a biotechnological tool to increase cucurbitacins in several genera of this family. Additionally, overexpression of a novel acyltransferase from cucurbitacin producing Iberis amara generates a hitherto undescribed acetylation at the C3-hydroxyl group of the cucurbitadienol backbone. While overexpression of the cytochromes P450 CsCYP88L2 and McCYP88L7 from Cucumis sativus and Momordica charantia (respectively), results in accumulation of new spectral feature as revealed by High resolution liquid chromatography mass spectroscopy analysis; the m/z of the new peak supports it might be a cucurbitacin hydroxylated at the C19 position in C. pepo hairy roots. Finally, this paper is a case study of how hairy roots can be used to metabolically engineer and introduce novel modifications in metabolic pathways that have not been fully elucidated.